Multiple physiological processes (i.e., breathing, blood flow, cerebrospinal fluid and aqueous humor formation, and sperm activation) are modulated by carbon dioxide and/or bicarbonate. The only known mammalian signaling protein modulated by bicarbonate or carbon dioxide is the bicarbonate responsive 'soluble'adenylyl cyclase (sAC). Mammalian sAC enzymes are molecularly, biochemically, and structurally distinct from the G protein regulated transmembrane adenylyl cyclases which mediate cellular responses to hormones. With previous support from this grant, we have confirmed that sAC serves as a cellular sensor for extracellular bicarbonate in sperm and as a cellular sensor for extracellular pH (via fluctuations of intracellular bicarbonate) in the epidydymis. We now propose to continue this project by testing whether sAC serves as the bicarbonate and/or carbon dioxide sensor in other physiological processes. In preliminary data for this application we show that somatic cells express isoforms of bicarbonate responsive sAC distinct from those previously identified in male germ cells. We plan to: (1) purify the somatic forms of sAC from bicarbonate responsive tissues and cell lines and determine their peptide amino acid sequences. (2) Clone these novel sAC isoforms. (3) Characterize their biochemical properties using both the native enzymes and heterologously expressed, recombinant sAC isoforms. (4) Ascertain whether and which sAC isoforms are expressed in various bicarbonate/carbon dioxide regulated physiological systems.